The Luteinizing Hormone Receptor (LHR): Characterization of regulatory mechanisms revealed that the human LHR (hLHR) expression is subject to regulation by a complex network and the participation of various signaling cascades through its promoter region. The histone deacetylase HDAC1/2/mSin3A complex was shown to be recruited to the proximal Sp1 binding site to elicit chromatin-localized condensation and silencing of the LHR gene. The status of DNA methylation and histone acetylation in the LHR promoter region operates coordinately to orchestrate the LHR level. PI3K/PKCzeta signaling pathway was shown to be critical for trichostatin A (TSA)-induced LHR transcriptional activation through phosphorylation of Sp1 at Ser641 and release of p107, which interacts with Sp1. LRH transcriptional activation is initiated by changes in chromatin structure induced by TSA that causes the release of a phosphatase(PP1 or PP2A) which is associated with Sp1 at the promoter. Other studies also demonstrated a critical role of the PKCalpha/Erk cascade in PMA-induced activation of LHR gene through Sp1 phosphorylation (other than Ser641 site), which causes release of the HDAC1/mSin3A repressor complex anchored at Sp1 sites. The phosphorylation induced by PKCalpha/ERK signaling disrupts the interaction between Sp1 and HDAC1 not HDAC2 and this causes release of HDAC1/mSin3A complex to activate hLHR. This is consistent with our finding that mSin3A functions as corepressor for HDAC1 but not HDAC2 in the regulation of LHR gene expression. Following the exclusion of a number of coactivators on Sp1-directed basal and TSA- induced transcriptional activation including p300, CBP, PCAF and others, we investigated the participation of PC4, a potential coactivator of Sp1, in the transcriptional control of LHR expression. Knockdown of PC4 expression reduced basal and TSA-induced LHR promoter activity and gene expression in MCF-7 cells. PC4 synergistically enhanced Sp1-but not Sp3- mediated LHR transcriptional activity. PC4 directly interacted with Sp1 at the LHR promoter and this interaction is negatively regulated by PC4 phosphorylation. PC4 associated with the LHR promoter basally and further recruitment was observed upon TSA treatment. TSA-induced recruitment of TFIIB and RNAP II was prevented by abolition of PC4 expression. PC4 function(s) in LHR transcription are beyond and independent of TSA-induced release of phosphatases from Sp1, Sp1 phosphorylation, and HDAC Sin3A co-repressor release indicating his role as linker coactivator of Sp1 and the transcriptional machinery. Gonadotropin regulated genes: 1)Gonadotropin-regulated long chain fatty acid Acyl (GR-LACS), identified in our laboratory, is a member of the LACS family regulated by LH/hCG in the rat Leydig cell (LC). Its mouse/human homologs, lipidosin/bubblegum, have been suggested by others to participate in X-linked adrenoleukodystrophy (X-ALD), a disorder with accumulation of very Long Chain FA (VLCFA) in tissues and blood. A GRLACS/lipidosin null mice generated in our laboratory did not support its association with X-ALD. 2) Gonadotropin Regulated Testicular Helicase (GRTH/Ddx25), a multifunctional enzyme discovered in our laboratory, is an essential regulator of sperm maturation. GRTH null mice are sterile due to spermatid arrest and failure to elongate. Through its association with CRM-1, GRTH participates in the export of selected messages relevant to the progress of spermatogenesis from the nuclear to cytoplamic sites, including the Chromatoid Body (CB) of spermatids (equivalent to somatic P bodies, contains members of RISC-complex and is viewed as a storage/processing site of mRNAs during spermatogenesis) and polyribosomes where it participates in translation of proteins that are essential for spermatogenesis. GRTH was found to also transport its own message to cytoplasmic sites. Our studies investigated thhe nuclear/cytoplasmic shuttling of GRTH in germ cells and its impact on the structure of the CB of spermatocytes in culture using immunofluorescence and EM studies. GRTH resides in the nucleus, the cytoplasm and the CB. Treatment of these cells with an inhibitor of nuclear export caused nuclear retention of GRTH and its absence in the cytoplasm and CB. Proteins of the RISC complex that do not participate in mRNA transport including MIWI and MVH and reside in the CB and cytoplasm were excluded from the CB and accumulated in the cytoplasm upon treatment with the inhibitor. This also occurred in spermatids of KO mice. The CB is changed from a large lobular-filamentous to a small condensed structure after treatment, resembling the CB of the GRTH-KO. GRTH did not interact with RISC members. Thus, rather than participating in translation regulation (either repression or degradation) via the small RNA pathway in the CB, the nuclear export of GRTH-RNP complexes could be required to maintain CB structural and functional integrity and to deliver essential RNAs presumably for silencing/storage/and or degradation of genes during germ cell development. Also, GRTH could be involved in the transport of mRNAs from CB to the polysomes for translation. GRTH is regulated by LH via cAMP and androgen at the transcriptional level directly in Leydig cells (LC) in vivo and in vitro and presumably indirectly in germ cells via genes induced by AR in Sertoli cells. Using transgenic mice we concluded that expression of GRTH in LC is directed by 205 bp minimal promoter activity and that the 5'1 kb fragment contains an non-clasical AR for androgen regulation of its expression in LC. Prolactin receptor (PRLR): -Structure function: Our early studies demonstrated 1) the presence of long (LF) and short forms (SF) of PRL;2) the formation of homo- and heterodimers of these forms in the absence of hormone and the action of PRL as a conformational modifier that induces activation of the JAK2/STAT5 signaling through LF and of JAK2 signalling through SF;3) The short form of the PRLR (S1b), with a short cytoplasmic tail silences PRL-induced activation of gene transcription by the long-form. Mutation of any one of the two paired Cys in S1b (S1bx)in extracellular subdomain 1- D1) eliminated the inhibitory action of S1b. The constitutive JAK2 phosphorylation and association observed in S1b was not present in cells expressing S1bx. Computer modeling based on PRLR crystal structure (extracellular) showed that minor changes in the tertiary structure of D1 upon S-S disruption propagated to the quaternary structure of the homodimer, affecting the dimerization interface (D2). Wild- type dimers were stabilized only through H-bonds in D2 subdomains, while the mutant developed additional H-bonds that bridged D1s. This explains the higher homodimerization affinity of the mutant (S1bx) revealed in BRET studies and provide the structural basis for its lack of inhibitory function. The PRLR conformation as stabilized by S-S bonds is required for the inhibitory action of S1b on PRL-induced LF-mediated function and JAK2 association/activity. -Transcriptional studies: Transcription of the human PRLR is governed by an E2/ERa mechanism independent of a E2 responsive element. E2/ERa through complex formation with SP1 and C/EBPb that associate to cognate elements leads to coactivator assembly and recruitment of TFIIB and Pol II. The basic region and leucine zipper of c/EBPbeta,zinc finger motifs of SP1 and DNA binding domain of ERa are the regions responsible for interaction. BRET assays further demonstrated interactions of ERa/ERa ERa/C/EBPb,ERa/SP1 which were increased by E2. Also, interactions of C/EBPb/SP1, C/EBPb/C/EBPb, SP1/SP1 were observed. BRET complementation assay revealed ERa constituitive dimers whose conformation induced by E2 favours its association with C/EBPb and SP1 mono- and/or homodimers.